The perception of sound and balance depends on the mechanosensory function of the hair bundle, a group of actin-filled stereocilia that project from the apical surface of auditory and vestibular hair cells. The study of deafness-linked genes has led to the identification of a number of proteins that are localized to the hair bundle and contribute to its development and function [1]. Despite these advances, the complete network of proteins that constitute the hair bundle and regulate its maturation, maintenance and function remain poorly defined. Although the mechanisms by which proteins are specifically transported to the bundle are also poorly understood, accumulated evidence suggests that myosin motor proteins may play important roles [2]. The goal of this proposal is to identify the bundle proteins that are transported to and maintained within the hair bundle during its formation and to test the hypothesis that myosin-VIIa (MYO7A) plays a role in the transport of these proteins. Using proteomics approaches as well as other biochemical techniques, the aims in this proposal will 1) identify in an unbiased manner the protein networks that are enriched within the hair bundle during and after bundle assembly, 2) determine whether MYO7A plays a role in the transport of specific proteins to the bundle and 3) identify which of these bundle-specific proteins directly or indirectly interact with MYO7A. The results of these studies will significantly contribute to our molecular understanding of bundle assembly and function and could help guide the development of therapeutic strategies for hair cell repair and regeneration.